The present invention relates to an antifreezing agent used in an aqueous medium system, comprising a protein which is enzymatically mcdified with covalent incorporation of an amino acid ester.
Preservation of aqueous medium systems in foods, biological bodies, and the like has been performed, in most cases, at a low temperature or in frozen state. When the aqueous medium systems mentioned above are subjected to a temperature below the freezing point of the aqueous system, however, tissue destruction results because of formation of ice crystals. Such destruction takes place by shearing the tissue with a knife of the formed ice crystal. Size of ice crystal growth maximizes at a temperature of -1.degree. to -5.degree. C.
There have been made various attempts in order to prevent the above-mentioned type of destruction of tissues. For example, in order to prevent ice crystals from growing to huge crystals, there is practiced a method in which the aqueous medium system is rapidly cooled to a temperature of -30.degree. C. to -50.degree. C. by using liquid nitrogen, whereby the temperature of the system can quickly pass through the temperature zone of maximum ice crystal formation. According to this method, the formed ice crystals are fine. There is also practicized a method in which to the aqueous medium system is added an antifreezing agent such as polyvinyl pyrrolidone which makes the freezing of the system in the temperature zone of maximum ice crystal formation inhibit.
However, the rapid freezing method described as the former method implies defects that it is unavoidable to lower the quality of products to a certain extent, and moreover that application of the method is restricted from a viewpoint of equipment requirement, as well as the consumption of a tremendous amount of energy for freezing the raw materials, and for storing and thawing the products. The latter method in which an antifreezing agent is added to the system also implies defects that all agents known as antifreezing agents cannot be used for foods, and that the desired antifreezing activity cannot be obtained unless the antifreezing agents are added in a relatively large amount.
On the other hand, in order to endow a protein with various functions, there is proposed a method in which verious chemical species are covalently attached to proteins in the presence of an enzyme. In many cases, the enzymatically modified protein (hereinafter referred to as "EMP") is considerably differ in function from the original protein.
In the course of studying on EMP, the present inventors have found the fact that an amino acid attached EMP prepared by using an amino acid ester as a chemical species exhibits an excellent antifreezing effect in various aqueous medium systems.
An object of the present invention is to provide an antifreezing agent used in an aqueous medium system, which is not only safe in biological bodies and effective in a small amount, but also able to maintain a stable unfrozen state over a long period of time.